Project Summary: Thyroid hormone (T3) regulates growth, development, and metabolic rate. Thyroxine (T4) must be activated by the type 1 and 2 deiodinases (D1 and D2) to produce T3. D1 and the type 3 deiodinase (D3) can also inactivate T3 by removal of an additional iodine. Thus, these enzymes control T3 availability. The goal of this proposal is to understand the interplay of these enzymes in context of each other and the central thyroidal axis such that situational-appropriate thyroid hormone concentrations and receptor occupancy are maintained. To further understand these networks, we created the C3H-D2KO mouse where the targeted disruption of the Dio2 gene is backcrossed into C3H mice with genetically low D1 (1/10 normal levels). Surprisingly, these mice maintain normal serum T3 levels, although T4 and TSH are increased. In Specific Aim 1 we will evaluate the tissue-specific thyroid status of these mice, define the mechanisms they use to achieve normal serum T3 concentrations, and evaluate their ability to compensate when challenged by a low iodine diet. In Specific Aim 2 we will investigate the causes of a 30% increase in body fat, and evaluate their response to a high fat diet. In Specific Aim 3, we will isolate D3 interacting proteins using 2 dimensional differential gel electrophoresis combined with mass spectrophotometry (2-D DICE MS) and the yeast two-hybrid system, with the goal of linking D3 cell biology to physiology. Further, we will assess if the first D3 interacting protein identified by 2-D DICE MS acts as a reducing co-factor for D3. Relevance to NIDDK Mission: This proposal will support basic research related to the fields of endocrinology, energy balance, development and metabolism. Relevance to Public Health: Thyroid hormone (T3) is necessary for proper fetal and neo-natal growth, while in adults it is a key regulator of metabolism and energy expenditure. This research will study how changes in levels of the enzymes that activate and inactivate T3 allow the correct amount of T3 to be present under a variety of different environmental conditions. Understanding how this balance is maintained is relevant to the health of the estimated 1-billion people living in areas of chronic iodine deficiency, and is important in identifying potential causes and treatments for the current obesity epidemic and the related diseases of diabetes, and heart disease.